Nucleic acid sequencing is one of the most widely used tools in molecular biology. Development of rapid and sensitive sequencing methods utilizing automated DNA sequencers has revolutionized modern molecular biology. In particular, analysis of entire genomes of plants, viruses, bacteria, fungi, and animals is now possible with next-generation sequencing technology.
Next-generation sequencing technology has brought a high level of efficiency to the process of genome sequencing. However, despite technology advances, whole genome sequencing is still associated with huge cost and workload since the related work flows tend to be complex, time-consuming, and costly to perform. In addition, there are various technical problems for accurate sample preparation, amplification and sequencing. As a result, the goal of rapid sequencing of a genome in a short period of time has not been possible.
Next-generation sequencing carried out in connection with target enrichment of certain types of genomic DNA sequencers (e.g., exons) allows one to focus on those targets. With this targeted next-generation sequencing variation, only the genomic areas of interest are sequenced, creating process efficiencies that reduce costs and allow more samples to be analyzed per study. Reducing the amount of DNA being interrogated allows investigators to perform the experiments with more statistically relevant numbers of samples.
Various approaches for targeted enrichment are available in the art. Most commonly used techniques are based on hybrid capture, PCR, and molecular inversion probes. For large target regions, hybrid capture has been the most efficient. A main advantage of this approach is enrichment in-solution rather than on microarrays; this provides easier handling and requires less DNA. In-solution capture, such as the SureSelect Target Enrichment System™ by Agilent Technologies, Inc., often applies biotinylated RNA bait molecules transcribed from DNA template oligo libraries, which are the key components and main costs. See US 20100029498, Mertes et al., Brief Funct Genomics 10:374-386, Gnirke et al., Nat. Biotechnol. 27:182-189, and Albert et al., Nat. Methods 4:903-905.
However, targeted next-generation sequencing technology is very time-consuming. A major bottleneck and speed-limiting step for the successful implementation of the targeted next-generation sequencing technology has been in the front-end steps to selectively capture and enrich targeted exons or intron regions scattered over the genomic, mitochondria and other forms of DNA, in a rapid and cost-effective manner. Indeed, the current selective hybrid capture procedure often ranges from 16 hours to over 70 hours. See, e.g., US 20100029498 and the SureSelect Target Enrichment System™ by Agilent Technologies, Inc. As a result, it usually takes as long as 2-4 days to generate a sample that is ready to be sequenced and even loner time to complete an entire sequencing process.
Thus, there is a need for compositions and methods of rapid target enrichment or selection of nucleic acids for next generation sequencing.